Chronic pain is a neurological disorder that impacts the lives of millions of Americans. Current treatments for chronic pain are limited by abuse potential and intolerable side effects. Endogenous proteases contribute to acute and chronic pain through the direct activation of the protease activated receptor-2 (PAR2) G-protein coupled receptor (GPCR). PAR2 is known to play an important role in chemical, inflammatory and cancer-induced pain but the possible efficacy of PAR2 antagonists in these preclinical models has not been assessed due to lack of available tools or clinical candidate compounds. Moreover, activation of PAR2 can lead to engagement of multiple signaling pathways yet agonists/antagonists with signaling pathway specific efficacy have not been explored as potential tools for understanding the role of PAR2 signaling in nociception. Studies assessing PAR2 activation and signaling pathway specific efficacy in the peripheral nervous system are important for the potential clinical development of PAR2 ligands for the treatment of pathological pain in humans. We are developing novel ligands to PAR2 in an effort to better elucidate the role of this receptor in nociception and to develop compounds that may have clinical utility for the treatment of pain. The central hypothesis of this application is that PAR2 plays a pivotal role in causing acute pain and promoting chronic pain and that high affinity ligands of the PAR2 receptor will represent a novel class of analgesics with utility in a number of chronic pain conditions. Thus, the primary objectives of this proposal are to develop novel and specific ligands to PAR2, to fully elucidate PAR2 contribution to acute and chronic pain, and to evaluate PAR2 ligand efficacy as novel analgesics in preclinical pain models. Successful completion of these studies will result in: 1) the discovery and development of novel agonists/antagonists for PAR2; 2) determination of signaling pathways that are engaged or attenuated by these novel agonists/antagonists; and 3) in vivo validation of PAR2 as an important pharmacological target for pain treatment. From these studies we intend to gain a more complete understanding of the physiological ramifications of different modes of agonist action at PAR2 as it relates to the nociceptive system. Successful studies will provide a preclinical rationale for the further development and testing of PAR2 ligands for the treatment of pain. Such findings represent a major step forward in the development of novel ligands to treat acute/chronic pain that can result from a variety of pathologies in humans.